Abstract:

Arbuscular mycorrhizal fungi (AMF) are the most prominent mycobionts of plants in the tropics, yet little is known about their diversity, species compositions and factors driving AMF distribution patterns. To investigate whether elevation and associated vegetation type affect species composition, we sampled 646 mycorrhizal samples in locations between 1000 and 4000 m above sea level (masl) in the South of Ecuador. We estimated diversity, distribution and species compositions of AMF by cloning and Sanger sequencing the 18S rDNA (the section between AML1 and AML2) and subsequent derivation of fungal OTUs based on 99% sequence similarity. In addition, we analyzed the phylogenetic structure of the sites by computing the mean pairwise distance (MPD) and the mean nearest taxon difference (MNTD) for each elevation level. It revealed that AMF species compositions at 1000 and 2000 masl differ from 3000 and 4000 masl. Lower elevations (1000 and 2000 masl) were dominated by members of Glomeraceae, whereas Acaulosporaceae were more abundant in higher elevations (3000 and 4000 masl). Ordination of OTUs with respect to study sites revealed a correlation to elevation with a continuous turnover of species from lower to higher elevations. Most of the abundant OTUs are not endemic to South Ecuador. We also found a high proportion of rare OTUs at all elevations: 79–85% of OTUs occurred in less than 5% of the samples. Phylogenetic community analysis indicated clustering and evenness for most elevation levels indicating that both, stochastic processes and habitat filtering are driving factors of AMF community compositions.

Abstract:

Orchid mycorrhizal detection, based only on fungal isolation from roots, is biased due to difficulties in isolating the respec- tive fungi. Previous investigations have shown that mostly ascomycetes, housed in the sheltered compartment of the vela- men covering the roots of epiphytic orchids, are isolated on agar plates. Roots of 83 individual epiphytic orchid of Stelis hallii, S. superbiens, S. concinna, Stelis sp. and Pleurothallis lilijae were sampled in the Reserva Biológica San Francisco, Southern Ecuador. The velamen was partly removed and root tissue macerated and transferred to Petri dishes containing corn meal agar, malt extract agar and Melin Norkrans modified medium. DNA was extracted from the cultures and the 5.8S-ITS region and partial nrLSU sequenced. Phylogenetic analysis revealed members of 12 orders of ascomycetes associ- ated with the roots of these orchids, a much higher diversity than previously known. Twelve isolates were related to the Helotiales. Our results have uncovered some of the fungal diversity within the velamen of epiphytic orchids; fungi invading, most likely, from tree bark or humus accumulation close to the roots.

Abstract:

Delimitation of species and the search for a proper threshold for defining phylogenetic species in fungi are under discussion. In this study, morpho- logical and molecular data are correlated to delimit species of Tulasnella, the most important mycobionts of Orchidaceae, which suffer from poor taxonomy. Resupinate basidiomata of Tulasnella species were collected in Ecuador and Germany, and 11 specimens (seven from Ecuador, four from Germany) were assigned to traditional species concepts by use of morphological keys. The specimens were compared by micro-anatomical examination with 75 specimens of Tulasnella borrowed from fungaria to obtain better insights on variation of characters. Sequences of the ITS region (127) were obtained after cloning from the fresh basidiomata and from pure cultures. Proportional variability of ITS sequences was analyzed within and among the cultures and the specimens designated to different morphospecies. Results sug- gested an intragenomic variation of less than 2%, an intraspecific variation of up to 4% and an interspe- cific divergence of more than 9% in Tulasnella. Cryptic species in Tulasnella, mostly from Ecuador, were revealed by phylogenetic analyses with 4% intraspecific divergence as a minimum threshold for delimiting species. Conventional diagnostic morpho- logical characters appeared insufficient for species characterization. Arguments are presented for molec- ular delimitation of the established species Tulasnella
albida, T. asymmetrica, T. eichleriana, T. cf. pinicola, T. tomaculum and T. violea.

Abstract:

Arbuscular mycorrhizae are important for growth and survival of tropical trees. We studied the community of arbuscular mycorrhizal fungi in a tropical mountain rain forest and in neighbouring reforestation plots in the area of Reserva Biológica San Francisco (South Ecuador). The arbuscular mycorrhizal fungi were analysed with molecular methods sequencing part of the 18S rDNA. The sequences were classified as Operational Taxonomic Units (OTUs). We found high fungal species richness with OTUs belonging to Glomerales, Diversisporales and Archaeosporales. Despite intensive sampling, the rarefaction curves are still unsaturated for the pristine forest and the reforestation plots. The communities consisted of few frequent and many rare species. No specific interactions are recognizable. The plant individuals are associated with one to ten arbuscular mycorrhizal fungi and mostly with one to four. The fungal compositions associated with single plant individuals show a great variability and variety within one plant species. Planted and naturally occurring plants show high similarities in their fungal communities. Pristine forest and reforestation plots showed similar richness, similar diversity and a significantly nested structure of plant-AMF community. The results indicate that small-scale fragmentation presently found in this area has not destroyed the natural AMF community, at least yet. Thus, the regeneration potential of natural forest vegetation at the tested sites is not inhibited by a lack of appropriate mycobionts.

Abstract:

Fungi are believed to be diverse in the tropics, but because many groups are
only known from their DNA sequences this hampers comparative diversity studies. We
investigated mycorrhizal Sebacinales (Basidiomycota) of 67 individuals of Ericaceae and
Orchidaceae in a tropical mountain ecosystem in Southern Ecuador to provide a ?rst
estimate of whether these fungi are particularly diverse in the Northern Andes. We par-
tially sequenced the internal transcribed spacer (ITS) and large subunit (LSU) regions of
the nuclear ribosomal DNA and analyzed them together with all Sebacinales sequences
available from GenBank. The clustering optimization technique was used to determine
clustering parameters that maximize the comparability between molecular operational
taxonomic units (MOTUs) obtained from the distinct loci. Sampling effort and species
richness were estimated with rarefaction-accumulation curves and non-parametric esti-
mation using Chao2 and compared between Southern Ecuador and France. Clustering
optimization indicated that a 1% LSU distance threshold corresponds to the commonly
used 3% dissimilarity threshold for ITS, and that a clustering algorithm close to single-
linkage clustering is optimal. The resulting clusters show that about 8?9% of observed Sebacinales MOTUs occur in the study area and that most of these MOTUs are endemic
(74%). The widespread MOTUs from Southern Ecuador were also found in Panama, North
America and Europe. The estimation of species richness revealed unsaturated sampling of
Sebacinales in general and also in our study area. Our results suggest a high diversity of
Sebacinales associated with Ericaceae and Orchidaceae at the study site in Southern
Ecuador, but no hotspot of Sebacinales in comparison with other areas.

Abstract:

Neotropical Vaccinioideae (Ericaceae) are evolutionary rather young and presumably of Northern Hemisphere origin. Vaccinioideae are highly dependent on their mycorrhizal symbionts and Sebacinales (basidiomycetes) were previously found to be the dominant mycobionts of Andean Clade Vaccinioideae (Neotropical Vaccinieae). We were interested to see whether the North American Vaccinioideae reached the Neotropics with their mycobionts or whether they acquired new, local Sebacinales.
We investigated Sebacinales of 58 individuals of Vaccinioideae from Ecuador, Panama and North America to examine whether mycobionts of each region are distantly or closely related.
We isolated the ITS of the ribosomal nuclear DNA in order to infer a molecular phylogeny of Sebacinales and to determine Molecular Operational Taxonomic Units (MOTUs). MOTU delimitation was based on a 3% threshold of ITS variability and conducted with complete linkage clustering. The analyses revealed that most Sebacinales from Ecuador, Panama and North America are closely related and that two MOTUs out of 33 have a distribution ranging from the Neotropics to the Pacific Northwest of North America. The data suggest that Neotropical and temperate Vaccinioideae of North America share their Sebacinales communities and that plants and fungi migrated together.

Abstract:

Mycorrhizas of vascular plants and mycorrhiza-like associations of liverworts are integral parts of terrestrial ecosystems, but have rarely been studied in tropical mountain rain forests. Our lightand electron microscopical studies at the RBSF revealed that the roots of nearly all tree species were well colonized by structurally diverse arbuscular mycorrhizal Glomeromycota (Haug et al. 2004, Beck et al. 2005), a few tree species by ectomycorrhizal Ascoand Basidiomycota (Haug et al. 2005), and that epiphytic, pleurothallid orchids formed mycorrhizas with Tulasnella species and members of the Sebacinales (Basidiomycota) (Suárez et al. 2006, Kottke et al. 2007). Species of Sebacinales also occurred in mycorrhizas of hemiepiphytic ericads (Setaro et al. 2006) and Tulasnella species were found in liverworts belonging to the Aneuraceae (Kottke et al. 2007). Traditionally, studies on biodiversity and host specificity have been based on morphologically defined species. No such approach was feasible in the case of the mycobionts in our study as the fungi did not display sufficient structural differences in the mycorrhizas for delimitation of morphospecies. The identification of the mycorrhiza-forming fungi in the forest could be done neither by spore nor by fruiting-body sampling, as both methods would have yielded only a very narrow spectrum of the fungal communities (Husband et al. 2002, Sanders 2004a). Instead, direct sequencing of the associated fungi from the plant material was carried out (Kottke et al. 2007). Given our present stage of knowledge, the sequence types (ribosomal genotypes) can rarely be precisely related to either morphological or biological species. However, the amount of information derived from the sequences of the ribosomal genes appeared to be meaningful in previous ecological studies on arbuscular mycorrhizas (Helgason et al. 2002, Husband et al. 2002, Sanders 2004b) as well as on mycorrhiza-forming Basidiomycota (Bidartondo et al. 2003, Bidartondo et al. 2004). However, analysis of biodiversity and specificity of the mycobionts from field samples using DNA sequences also poses problems. Firstly, results are limited by the available primers. Secondly, problems resulted from the observation that the ribosomal genes can show intraspecific variation, especially in the case of the multinucleate Glomeromycota (Sanders et al. 1995, Lloyd-Macgilp et al. 1996, Clapp et al. 2001, Sanders 2004b). These facts pose general, unresolved challenges to a species concept based on meaningful levels of genetic diversity. Thus the list of fungal sequences presented here is far from being complete and far from being a species list.

Abstract:

Abstract: For the first time in tropical mountain rain forest, arbuscular mycorrhizal fungal
richness and community composition was investigated from planted seedlings of Cedrela
montana, Heliocarpus americanus, Juglans neotropica and Tabebuia chrysantha in reforestation
plots on degraded pastures. A segment of fungal 18S rDNA was sequenced from the mycorrhizas.
Sequences were compared with those obtained from mycorrhizas of adult trees of 30
species in the neighboring, pristine tropical mountain rain forest. In total, 193 glomeromycotan
sequences were analyzed, 130 of them previously unpublished. Members of Glomeraceae,
Acaulosporaceae, Gigasporaceae and Archaeosporales were found in both habitats, with Glomus
Group A sequences being by far the most diverse and abundant. Glomus Group A sequence type
richness did not appear to differ between the habitats; a large number was observed in both.
Glomus Group A sequence type composition, however, was found distinctly different. Seedlings
were rarely colonized by fungi of the pristine forest but trapped a number of fungi known from
other areas, which were rarely found in the pristine forest.

Abstract:

Distinctive groups of fungi are involved in the diverse mycorrhizal associations of land plants. All previously known mycorrhiza forming Basidiomycota associated with trees, ericads, liverworts or orchids are hosted in Agaricomycetes, Agaricomycotina. Here we demonstrate for the first time that Atractiellomycetes, members of the ?rust? lineage (Pucciniomycotina), are mycobionts of orchids. The mycobionts of 103 terrestrial and epiphytic orchid individuals, sampled in the tropical mountain rain forest of Southern Ecuador, were identified by sequencing the whole ITS1-5.8S-ITS2 region and part of 28S rDNA. Mycorrhizae of 13 orchid individuals were investigated by transmission electron microscopy. Simple septal pores and symplechosomes in the hyphal coils of mycorrhizae from four orchid individuals indicated members of Atractiellomycetes. Molecular phylogeny of sequences from mycobionts of 32 orchid individuals out of 103 samples confirmed Atractiellomycetes and the placement in Pucciniomycotina, previously known to comprise only parasitic and saprophytic fungi. Thus, our finding reveals these fungi, frequently associated to neotropical orchids, as the most basal living basidiomycetes involved in mycorrhizal associations of land plants.

Abstract:

Orchids depend on fungi for germination and protocorm development, but also form mycorrhizas during life time. Mycorrhizal fungi are, therefore, discussed as driving force in orchid evolution and speciation. However, for serious discussion reliable data on fungal identity are crucial. Application of molecular phylogeny and transmission electron microscopy congruently revealed distinct fungal groups as orchid mycobionts. Sebacinales Group B, Tulasnellales, and Ceratobasidiales were found associated with terrestrial orchids in open grasslands and arbuscular mycorrhizal forests and with epiphytes, while Sebacinales Group A, Thelephorales, Russulales, some Euagaricales and Tuberales form mycorrhizas with terrestrial orchids in ectomycorrhizal forests. Enzyme and isotope analyses revealed that the former obtain carbon from rotten organic material to nourish the protocorm while the latter take carbon from ectomycorrhizas of woody plants to feed protocorms and adult heterotrophic or mixotrophic plants. Mycobionts of terrestrial orchids appeared to be of narrower host range than previously expected, and co-speciation was discussed. The few investigations on mycobionts of epiphytic orchids so far indicated sharing of hosts. Further information is needed of mycobionts from tropical terrestrial and epiphytic orchids at well resolved molecular level of fungal identity, in situ prove, host range, and ecology to finally evaluate if association strategies differ between epiphytic and terrestrial orchids or between temperate and tropical habitats.